Catalytic antibody technology can often provide a rapid and versatile entry into new catalytic proteins. (R. A. Lerner, et al., Science (1991): vol. 252, pp 659-667; and P. G. Schultz, et al., Acc. Chem. Res. (1993), vol. 26, pp 391.) A number of enzymatic processes have been successfully mimicked by catalytic antibodies. Tramontano and others have shown that catalytic antibody can be generated which have hydrolase activity with respect to ester bonds. (A. Tramontano et al., J. Am. Chem. Soc. (1988): vol. 110, pp 2282; K. D. Janda et al., Science (1989): vol. 244, pp 437; J. Guo et al., J. Am. Chem. Soc. (1994): vol. 116, pp 6062; and G. W. Zhou, et al., Science (1994): vol. 265, pp 1059.) Similarly, Janda and others have shown that catalytic antibody can be generated which have hydrolase activity with respect to amide bonds. (K. D. Janda et al., Science (1988): vol. 241, pp 1188; and M. T. Martin et al., J. Am. Chem. Soc. (1994): vol. 116, pp 6508.) Catalytic antibodies having hydrolytic activity with respect to a variety of glycosidic bonds have also been generated. (J. L. Reymond et al., Angew. Chem. Int. Ed. Engl. (1991): vol. 30, pp 1711; and J. Yu, et al. Angew. Chem. Int. Ed. Engl. (1994): vol. 33, pp 339.) Catalytic antibodies having the ability to form amide bonds have been generated by several investigators. (S. J. Benkovic et al., Proc. Natl. Acad. Sci. USA (1988): vol. 85, pp 5355; J. R. Jacobsen et al., Science (1992): vol. 256, pp 365; R. A. Gibbs et al. Science (1992): vol. 258, pp 803; ; R. Hirschmann et al., Science (1994): vol. 265, pp 234; and J. R. Jacobsen et al., Proc. Natl. Acad. Sci. USA (1994): vol. 91, pp 5888.) Decarboxylation reactions can be catalyzed using catalytic antibody generated for that purpose. (C. Lewis et al., Science (1991): vol. 253, pp 1019; J. A. Ashley et al., J. Am. Chem. Soc. (1993): vol. 115, pp 2515; and T. M. Tarasow et al., J. Am. Chem. Soc. (1994): vol. 116, pp 7959.) The heme-mediated reduction of hydrogen peroxide can be catalyzed using catalytic antibody (A. G. Cochran et al., J. Am. Chem. Soc. (1990): vol. 112, pp 9414.) The 3,3-sigmatropic rearrangement catalyzed by chorismate mutase can also be catalyzed by catalytic antibody. (D. Y. Jackson et al., J. Am. Chem. Soc. (1988): vol. 110, pp 4841; D. Hilvert et al., J. Am. Chem. Soc. (1988): vol. 110, pp 5593; D. Y. Jackson et al., Angew. Chem. Int. Ed. Engl. (1992): vol. 31, pp 182; and M. R. Haynes et al., Science (1994): vol. 263, pp 646.) The imitation of natural enzymes by catalytic antibodies is a continuing challenge as well as a source of inspiration in this field.
Many enzymes rely on imine and enamine intermediates to catalyze exchange reactions at the .alpha.-carbon of carbonyl compounds. Aldolases catalyze the aldol addition of ketones with aldehydes and decarboxylases catalyze the decarboxylation of .beta.-keto acids. The catalytic group in both enzymes is the .epsilon.-amino-group of a lysine residue. A related reaction is the interconversion of .alpha.-keto and .alpha.-amino acids catalyzed by transaminases using the cofactor pyridoxamine phosphate. The catalytic mechanisms of each of these reactions are reviewed by D. J. Hupe, Enzyme Mechanisms, Ed. M. I. Page and A. Williams, 1987, p. 317-344 and by W.-D. Fessner, Kontakte (1992): vol. 3, pp 3-9.
The mechanism of these reactions, illustrated below for aldolases (scheme 1), involves initial condensation of a carbonyl group of the substrate with the amine to form an iminium intermediate I. The iminium cation is the protonated form of the imine. Both forms are in rapid equilibrium and their relative proportions depends on the pH. The iminium is the kinetically relevant form. When employed herein, the term iminium generally refers to both forms. This intermediate then undergoes an exchange at the .alpha.-carbon of the carbonyl, a process which is facilitated by formation of enamine II. In the aldolase reaction, a proton is exchanged for an aldehyde, with formation of a new carbon-carbon bond. Finally, hydrolysis of the primary adduct liberates the carbonyl product and regenerates the free amino group, e.g., D. J. Hupe, in New Comprehensive Biochemistry, vol. 6, The Chemistry of Enzyme Action, Ed. M. I. Page, Elsevier, Amsterdam, 1984, chapter 8.
All of these reactions play central roles in biosynthetic pathways, including the biosynthesis, interconversion and degradation of sugars and amino-acids. The ability to mimic these processes using catalytic antibodies would offer significant opportunities for designing synthetically useful catalytic reactions. What is needed is a catalytic antibody directed to catalyzing aldol reactions using enamine chemistry, i.e., utilizing a primary amine as cofactor in a catalytic process as employed by aldolase enzymes. ##STR1##